Bradykinin (BK) is an endogenous nonapeptide (H-Arg-Pro-Pro-Gly-Phe-Ser-Pro-Phe-Arg-OH; SEQ ID NO: 1) that is generated by cleavage of the precursor polypeptide (kininogen) by specific proteases (kallikriens) within numerous tissues of the body (Regoli, D. and Barabe, J. Pharmacol. Rev., 32, 1-46, 1980; Hall, J. M., Pharmacol. Ther., 56, 131-190, 1992; Leeb-Lundberg et al., Pharmacol. Rev. 57: 27-77, 2005). Certain enzymes of the kininase family degrade BK and related peptides and thus inactivate these peptides. All components of the kallkrien/kinin system, including specific receptors activated by BK and related peptides, are present in the human eye cells and tissues (Ma et al., Exp. Eye Res. 63: 19-26, 1996; Sharif and Xu, Exp. Eye Res. 63: 631-637, 1996).
BK and another endogenous peptide (Lys-BK; Lys-Arg-Pro-Pro-Gly-Phe-Ser-Pro-Phe-Arg; SEQ ID NO: 2) interact with two major BK receptor-subtypes, namely B1 and B2 to produce their biological effects (Regoli, D. and Barabe, J. Pharmacol. Rev., 32, 1-46, 1980; Hall, J. M., Pharmacol. Ther., 56, 131-190, 1992; Leeb-Lundberg et al., Pharmacol. Rev. 57: 27-77, 2005). The B2-subtype is found under normal physiological conditions, while the B1-subtype is typically induced during injury or trauma (Hall, J. M., Pharmacol. Ther., 56, 131-190, 1992; Leeb-Lundberg et al., Pharmacol. Rev. 57: 27-77, 2005). While the B1-subtype has a low affinity for BK and a high affinity for Des-Arg9-BK (Arg-Pro-Pro-Gly-Phe-Ser-Pro-Phe; SEQ ID NO: 3) and Lys-BK, the B2-subtype has a high affinity for BK and Lys-BK but a low affinity for Des-Arg9-BK. Both receptor subtypes have been cloned and shown to be coupled to G-proteins and phospholipase C and their activation results in the generation of the second messengers inositol trisphosphate (IP3) and diacylglycerol (DAG) (Bhoola et al., Pharmacol. Rev. 44: 1080, 1992; Hall, J. M., Pharmacol. Ther., 56, 131-190, 1992; Leeb-Lundberg et al., Pharmacol. Rev. 57: 27-77, 2005). While IP3 mobilizes intracellular Ca2+ ([Ca2+]i), DAG phosphorylates protein kinase C, and together these events lead to the final biological response such as cell shape change, tissue contraction or fluid secretion or all of the above. Additional events ensuing from elevation of [Ca2+]i include activation of nitric oxide synthase (NOS) to produce nitric oxide (NO) that in turn activates guanylate cyclase to produce cGMP, and activation of cycloxygenases and/or phospholipase A2 that produce endogenous prostaglandins (PGs) that in turn elevate intracellular cAMP (Leeb-Lundberg et al., Pharmacol. Rev. 57: 27-77, 2005; Crider and Sharif, J. Ocular Pharmacol. Ther. 17: 59-67, 2001).
Activation of the B2-receptor can also lead to inhibition of cAMP production in host cells transfected with the human recombinant B2 receptors (Meini et al., Brit. J. Pharmacol. 143: 938-941, 2004). The majority of the physiological and pathological effects of BK are mediated by the B2-receptor-subtype. However, pharmacological evidence has pointed to two additional BK-receptor subtypes, namely B3 and B4 (Hall, J. M., Pharmacol. Ther., 56, 131-190, 1992; Sharma, Gen. Pharmacol., 24, 267-274, 1993). B3 and B4 receptor subtypes are actually stimulated by certain peptide BK antagonists whereas the B1 and B2 subtypes are blocked by the latter antagonists (Sharma, J. N., Gen. Pharmacol., 24, 267-274, 1993). While the presence of B3 or B4 receptor subtypes in the eye has not been investigated to date, there is precedence for their existence in this organ since there is a robust BK-precursor and BK-generating enzyme pool in human ocular tissues and the presence of B1 and B2 receptors (Ma et al., Exp. Eye Res., 63: 19-26, 1996).
Two new families of peptides related to BK, namely ovikinins (Schroder et al. J. Biol. Chem. 272: 12475-12481, 1997) and bombinakinins (Lai et al, Biochem. Biophys. Res Comm. 286: 259, 2001; Lai et al., Peptides, 24: 199, 2003; O'Rouke et al., Regul. Peptides 121: 65, 2004; Lee et al., Regul. Peptides, 127: 207, 2005), have been discovered recently that may interact with BK receptors or similar receptors, and which may be useful for lowering IOP. Additionally, a new receptor termed GPR100 has been recently discovered with which BK also interacts (Boels and Schaller, Br. J. Pharmacol. 140: 932-938, 2003). Likewise, other selective peptides of the dynorphin family (Lai et al. Nature Neurosci. 9: 1534-1540, 2006), neurotensin (Park and Kim, Cell. Signal., 15: 519-527, 2003) and neuropeptide Y (Gibbs et al., Br. J. Pharmacol. 150: 72-79, 2007), can activate BK receptors.
Additional useful properties imparted by BK or BK mimetics include the lowering of mRNA of connective tissue growth factor (CTGF) (Huang et al. Am. J. Physiol. Lung Cell Mol. Physiol. 290: L1291-L1299, 2006), a fibrotic cytokine that has been implicated in the possible etiology of ocular hypertension by promoting deposition of collagen and fibronectin in the TM area (International Patent Application No. PCT/US2003/012521 to Fleenor et al. published Nov. 13, 2003 as WO 03/092584 and assigned to Alcon, Inc.); BK-induced inactivation of RhoA (Am. J. Physiol. Lung Cell Mol. Physiol. 290: L129-L1299, 2006) since Rho kinase inhibitors lower ocular hypertension (Waki et al., Curr. Eye Res. 22: 470-474, 2001); BK-induced blunting of systemic hypertension (Majima et al., Hypertension 35: 437-442, 2000) and BK-induced increase in blood flow (Ito et al. Br. J. Pharmacol. 138: 225-233, 2003) that is beneficial for retinoprotection (Tamaki et al., J. Ocular Pharmacol. Ther. 15: 313-321, 1999). In addition, BK and its analogs would be anticipated to be useful therapeutically because BK attenuates the release of pro-inflammatory cytokines from activated microglial cells (Noda et al., J. Neurochem. 101: 397-410, 2007).
There are a number of ocular conditions that are caused by, or aggravated by, damage to the optic nerve head, degeneration of ocular tissues, and/or elevated IOP. For example, “glaucomas” are a group of debilitating eye diseases that are a leading cause of irreversible blindness in the United States and other developed nations. Primary Open Angle Glaucoma (“POAG”) is the most common form of glaucoma (Quigley, Br. J. Opthalmol., 80: 389-393, 1996). The disease is characterized by the degeneration of the trabecular meshwork, leading to obstruction of the normal ability of aqueous humor to leave the eye without closure of the space (e.g., the “angle”) between the iris and cornea (Rohen, Opthalmol. 90: 758-765, 1983; (Quigley, Br. J. Opthalmol., 80: 389-393, 1996). A characteristic of such obstruction in this disease is an increased IOP, resulting in progressive visual loss and blindness if not treated appropriately and in a timely fashion. The disease is estimated to affect between 0.4% and 3.3% of all adults over 40 years old. Moreover, the prevalence of the disease rises with age to over 6% of those 75 years or older. Thus, close to 70 million are afflicted by glaucoma (Quigley, Br. J. Opthalmol., 80: 389-393, 1996).
Glaucoma affects three separate tissues in the eye. The elevated IOP associated with POAG is due to morphological and biochemical changes in the trabecular meshwork (TM), a tissue located at the angle between the cornea and iris, and ciliary muscle (CM) bundles. Most of the nutritive aqueous humor exits the anterior segment of the eye through the TM. The progressive loss of TM cells and the build-up of extracellular debris in the TM of glaucomatous eyes leads to increased resistance to aqueous outflow, thereby raising IOP. Elevated IOP, as well as other factors such as ischemia, cause degenerative changes in the optic nerve head (ONH) leading to progressive “cupping” of the ONH and loss of retinal ganglion cells and axons. The detailed molecular mechanisms responsible for glaucomatous damage to the TM, ONH, and the retinal ganglion cells are unknown.
Twenty years ago, the interplay of ocular hypertension, ischemia and mechanical distortion of the optic nerve head were heavily debated as the major factors causing progression of visual field loss in glaucoma. Since then, other factors including excitotoxicity, nitric oxide, absence of vital neurotrophic factors, abnormal glial/neuronal interplay and genetics have been implicated in the degenerative disease process. The consideration of molecular genetics deserves some discussion insofar as it may ultimately define the mechanism of cell death, and provide for discrimination of the various forms of glaucoma. Within the past 10 years, over 15 different glaucoma genes have been mapped and 7 glaucoma genes identified. However, despite such progress, the glaucomas still remain poorly understood.
Glaucoma is a progressive disease which leads to optic nerve damage and, ultimately, total loss of vision. Since there is a good correlation between IOP control and prevention/reduction of glaucomatous damage in POAG patients (Mao et al., Am. J. Opthalmol. 111: 51-55, 1991), several therapeutic agents have been developed to treat ocular hypertension (Clark and Yorio, Nature Rev. Drug Discovery, 2: 448-459, 2003; Sharif and Klimko, Ophthalmic Agents, in Comprehensive Medicinal Chemistry II., Vol. 6, Chapter 6.12, p. 297-320; Eds: D. J. Triggle and J. B. Taylor, Elsevier Oxford, 2007). Thus, it is known that elevated IOP can be at least partially controlled by administering drugs which either reduce the production of aqueous humor within the eye, such as beta-blockers and carbonic anhydrase inhibitors, or increase the outflow of aqueous humor from the eye, such as miotics and sympathomimetics. Unfortunately, many of the drugs conventionally used to treat ocular hypertension have a variety of problems. For instance, miotics such as pilocarpine can cause blurring of vision and other visual side effects, which may lead either to decreased patient compliance or to termination of therapy. Systemically administered carbonic anhydrase inhibitors can also cause serious side effects such as nausea, dyspepsia, fatigue, and metabolic acidosis, which side effects can affect patient compliance and/or necessitate the termination of treatment. Another type of drug, beta-blockers, have increasingly become associated with serious pulmonary side effects attributable to their effects on beta-2 receptors in pulmonary tissue. Sympathomimetics, on the other hand, may cause tachycardia, arrhythmia and hypertension. Recently, certain prostaglandins and prostaglandin derivatives have been described in the art as being useful in reducing IOP. Typically, however, prostaglandin therapy for the treatment of elevated IOP is attended by undesirable side-effects, such as irritation and hyperemia of varying severity and duration. There is therefore a continuing need for therapies that control elevated IOP associated with glaucoma without the degree of undesirable side-effects attendant to most conventional therapies.